Modifying the mRNA (3 steps):

1.     Adding a 5’ cap

a.    After the synthesis of 20-30 nucleotides, the 5’ end of the mRNA is capped 5’ to 5’ with a guanine nucleotide

b.    A methyl group is added to 2’ OH

c.    This confuses the nuclease and prevents it from being degraded

d.    Cap binding proteins (CBPs) interact with the other proteins helping to stabilize the mRNA

2.    Splicing -> removing introns and joining exons (3 steps)

a.    Introns are non-coding while exons code for amino acids. These introns must be removed during RNA processing

b.    In order to locate the introns, **they always begin with 5’-GT (U) and end with AG-3’

c.    Cleavage occurs at the 5’ end which is free to join with an A via a phosphodiester bond in a specific site. This forms a lariat, which is later excised and the exons therefore are joined.

d.    Splicing is mediated by spliceosomes (snRNAs + proteins)

e.    These spliceosomes start to assemble during pre-mRNA synthesis

f.    **Clinical Correlate**

Mutation in pre-mRNA splicing factor causes Retinitis Pigmentosum. Mutation would prevent adequate splicing from occurring. Other diseases associated with mutations in genes involved in splicing are Spinal Muscular Atrophy, forms of cancer, trinucleotide repeat disorders, and myotonic dystrophy

3.       Polyadenylation

a.       This basically keeps the cap formed previously from being degraded by forming a loop with the help of poly(A) binding protein (PBP)

b.      Done at around 10-30 nucleotides of the 3’ end of the AAUAA A site

c.       Catalyzed by poly(A) polymerase